How Is TB Transmitted? Sébastien Gagneux, PhD 20 th March, 2008
Today s Outline 1) Global spread of Mtb Comparative genomics Phylogeny 2) Transmission of drug-resistant Mtb Fitness assays Molecular epidemiology
The Global Diversity of Mtb Global strain collection: 875 strains 80 countries TbD1 12 can 239 105 207 M. canettii Indo-Oceanic 181 East-Asian 9 150 142 750 East-African-Indian 122 Middle East 115 182 183 193 Americas Europe Large Sequence Polymorphisms pks 15/1 Δ7bp 219 H37Rv-like 174 726 West Africa Euro-American 761 South Africa 724 Central Africa 711 West-African-1 702 West-African-2 7, 8, 10 M. bovis lineage Gagneux et al. PNAS 2006
Geographic Distribution of Mtb Diversity Gagneux et al. PNAS 2006
Global TB Burden by Mtb Lineage Estimated number of prevalent cases in 2003 (WHO 2005) 6 5 4 3 Whole-genome sequences: H37Rv (Sanger) CDC1551 (TIGR) F11 (Broad) C (Broad) Haarlem (Broad) 210 (TIGR) 2 1 0 E & SE -Asia Indian sub-continent Sub-Saharan Africa C-Asia / Russia Americas Europe N-Africa Middle East Gagneux & Small Lancet ID 2007 Number of cases (millions)
Modern MTB Ancient MTB Global Phylogeny of M. tuberculosis 108 strains ~70kbp/strain 100 100 100 100 Animal adapted M. canettii 5 Gagneux et al. in preparation
Out-of-Africa of M. tuberculosis?
1) Global Spread of Mtb: Conclusions Origin of Mtb most likely in Africa Geographical distribution of Mtb diversity suggests Out-of-Africa But! More recent migration also involved (travel, trade, conquest) Different evolutionary histories might have resulted in lineage-specific phenotypic differences (vaccines!).
Where Is TB Now? Global TB 2005 Estimates All forms of TB MDR-TB Estimated number of cases 8.8 million 424,000 Estimated number of deaths 1.6 million 116,000 XDR-TB 27,000 16,000 >50% Mortality Entering post-antibiotic era!
The boundaries and names shown and the designations used on this map do not imply the expression of any opinion whatsoever on the part of the WHO concerning the legal status of any country, territory, city or area or of its authorities, or concerning the delimitation of its frontiers or boundaries. Dotted lines on maps represent approximate border lines for which there may not yet be full agreement. WHO 2005. All rights reserved Countries with confirmed XDR-TB Argentina Armenia Bangladesh Brazil Canada Chile China, Hong Kong SAR Czech Republic Ecuador France Portugal Georgia Republic of Korea Germany Russian Federation Islamic Republic of Iran Italy South Africa Japan Spain Latvia Sweden Mexico Thailand Norway Based on information provided to WHO Stop TB Department March 2007 UK Peru USA
XDR-TB in the
2) Transmission of Drug-resistant Mtb Public Health is important What about Biology? Is drug-resistance costly (to the bug)? Studies in E. coli suggest fitness cost MDR / XDR-TB associated with HIV Are XDR strains less fit?
Fitness: The Experimental Approach RIF S RIF R Conditioning Competition no RIF CFU measurements @ baseline & endpoint RIF
1 st strain background: CDC1551 CDC1551 RIF R mutants 200ul wildtype RIF 2 nd strain background: T85/Beijing T85 RIF R mutants 200ul wildtype RIF
Clinical Isolates with Acquired RIF R 4 to 37 months RIF S RIF R Same DNA fingerprint
Fitness Cost of Rifampicin-Resistant Mtb Lab-derived mutants: Mean relative fitness 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 S531L H526Y H526D S531W H526R S522L Q513L H526P R529Q rpob mutation Clinical strains: Mean relative fitness 1.3 1.2 1.1 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 1 2 3 4 5 6 7 8 9 10 S531L Isolate pair other rpob Gagneux et al. Science 2006
Clinical Frequency of rpob Mutations rpob mutation S531L H526Y H526D S531W H526R R529Q Mean fitness 1.02 0.82 0.78 0.82 0.82 0.58 Clinical frequency (%)* 54 11 7 4 3 0 * based on 840 clinical isolates (O Sullivan et al. 2005)
Fitness: The Molecular Epidemiology Approach DNA fingerprinting (IS6110 RFLP) reactivated transmitted
Population-based Molecular Epidemiological Study in San Francisco INH resistance caused by different mutations Different INH R mutations have different effects on bacterial virulence / fitness in animal models katg activates INH and is important for virulence Hypothesis: Mutants with high fitness cost will transmit less
Mutations in 152 INH R Isolates from SF (1991-1999) Mutation N (%) KatG activity 1) Non-functional KatG 34 (22.4) -- 2) katg S315T 62 (40.8) -+ 3) inha prom. -15 c t 39 (25.7) + + No mutation 17 (11.1) + + Gagneux et al. PLoS Pathogens 2006
INH R Mutation and RFLP Clustering Mutation KatG activity % RFLP clustering p-value 1) Non-functional KatG -- 0.0 reference 2) katg S315T -+ 11.3 < 0.05 3) inha -15 c t + + 17.8 < 0.01
The Biogeography of Mtb 12 can M. canettii 239 Indo-Oceanic TbD1 105 207 181 East-Asian 9 150 142 750 East-African-Indian 122 Middle East 115 182 183 193 Americas Europe pks 15/1 Δ7bp 219 H37Rv-like 174 West Africa Euro-American 726 761 South Africa 724 Central Africa 711 West-African-1 702 West-African-2 7, 8, 10 M. bovis lineage Gagneux et al. PNAS 2006
Does Strain Lineage Impact Propensity Towards Low / High-Cost INH R Mutations? Lineage / Mutation Blue Lineage: 1) Non-functional katg mutations Red Lineage: 2) katg S315T Pink Lineage: 3) inha prom. -15 c t Odds Ratio 5.6 2.0 3.8 P-value < 0.001 0.052 < 0.001
Blue Mtb (Beijing) Associated with MDR The Russia Gambia The Vietnam Gambia The South Gambia Africa
2) Transmission of Drug-resistant Mtb: Conclusions Fitness and transmission of drug-resistant Mtb is a function of: Specific mutation Strain genetic background Compensatory evolution (?) Implications for predicting the future of MDR / XDR-TB
Evolution of Drug Resistance No-cost mutation Pre-adapted genetic background Fitness DS DR DR DR Compensation Time
Acknowledgments ISB, Seattle Peter Small Hadar Sheffer Lee Rowen Jared Roach Marta Janer Scott Bloom NIMR, London Douglas Young Vivek Rao Damien Portevin Stanford Marcus Feldman Misha Lipatov Dmitri Petrov Ruth Hershberg Brendan Bohannan Alex Pym Clara Davis Long Gary Schoolnik Tran Van Funding: UC San Francisco Phil Hopewell Midori Kato-Maeda Swiss National Science Foundation Novartis Foundation National Institutes of Health Wellcome Trust